As cells divide the different cellular programs of cell division must be coordinated with one another. For example, the duplication of chromosomes, the separation of these duplicated chromosomes, and the division of one cell into two cells must all occur at precise times relative to one another and to other cellular events. To achieve this coordination, there are special times during the cell cycle, termed checkpoints, in which progression of the cell cycle can be paused. At these checkpoints, accurate/ undamaged completion of an earlier program is necessary before a later program will be initiated. Checkpoints work because the cell has specialized machines, made up of enzymes and other proteins, that are activated by damaged or incomplete cellular structures. Once activated these checkpoint machines cause the progression of the cell cycle to halt until the cellular structures are repaired and complete. When checkpoints are defective, cells continue through the cycle accumulating damaged structures and leading to disease states. Indeed, checkpoints are often bypassed in cancer cells, and defective checkpoints lead to higher risk of cancer. We recently proposed a novel checkpoint, here termed the ?Rlm1-dependent checkpoint?, that delays passage through the cell cycle under a particular stressful environmental condition. As the name suggests, this checkpoint was revealed by a deletion mutant in the gene encoding the Rlm1 transcription factor. Rlm1 is known to be activated in response to cell-wall stress, so it may be that this checkpoint responds to stress at the cell surface. Interestingly, bypassing the Rlm1 checkpoint, unlike bypassing known checkpoints, leads to cell death in only one of the two products of cell division. Our long term objective in this project is to characterize the function and mechanism of the Rlm1-dependent checkpoint. Our first specific aim is to characterize the position during the cell cycle at which this checkpoint operates, and the cell cycle regulators and additional cellular components through which it acts. Our second specific aim is to identify the nature of the signal to which this checkpoint responds and the nature of the damage that accumulates when the checkpoint is bypassed. To accomplish these aims we will employ a combination of genetic, cytological, and molecular biological assays. For example, we will grow both normal yeast and mutants defective in the Rlm1 checkpoint under non-stressful conditions and then release them suddenly into the stressful condition. We will then compare the normal and mutant yeast over time for known molecular or cellular events in the cell cycle as well as different types of cellular damage. We will test specific hypotheses regarding the mechanism of this checkpoint by examining mutants known to be defective in particular aspects of cell maintenance and cell cycle control for their role in this checkpoint. Characterization of this new type of checkpoint should reveal important new aspects of cell cycle control and may prove useful in understanding new aspects of disease states.